Agriculture Reference
In-Depth Information
to be dead. However, spores that have been killed in this fashion are actually not dead,
and they can be recovered if they are germinated artifi cially through degradation of
the spore cortex with exogenous lysozyme (Setlow and others 2002). This fi nding is
important to keep in mind when assessing spore killing by a particular agent. Are the
spores truly dead or only conditionally dead, and might they revive in alternative
recovery media or with treatment by an appropriate stimuli? This point is noted again
below.
Inner Membrane Damage
In addition to the mechanisms of spore inactivation considered above, a number of
chemicals that kill spores appear to damage the spore's inner membrane in some
fashion such that this membrane ruptures, either directly or upon spore germination
(Setlow 2006). One agent that has this effect is relatively concentrated mineral acids.
For example, incubation of
B. subtilis
spores in 1 M HCl rapidly causes dramatic
rupture of the dormant spore's inner membrane (Setlow and others 2002). Although
such a treatment is extremely effective in spore inactivation, clearly this method is
not likely to be used in the decontamination of foodstuffs.
There are a number of other agents that appear to kill spores by damaging the
spore's inner membrane and that are, or could be, used in the decontamination of fresh
produce and other foodstuffs. Agents that appear to kill spores by this mechanism
include chlorine dioxide, hypochlorite, ozone, and some oxidizing agents (Cortezzo
and others 2004; Genest and others 2002; Loshon and others 2001; Paul and others
2006, 2007; Young and Setlow 2003, 2004a,b). These agents do not kill spores by
DNA damage, and
-type SASP are not important in spore resistance to them. Upon
extended treatment with these agents, spores exhibit large decreases in germination
effi ciency. However, when spores are killed only 90-99%, the treated spores do still
germinate, albeit often slowly. In addition, artifi cial germination of partially killed
spore populations with lysozyme does not increase spore recovery (in contrast to the
situation with NaOH-treated spores described above). Thus, the reason for the killing
of spores by this group of agents is not the inactivation of one or more components
of the spore's germination apparatus.
There is evidence indicating that these three agents, and perhaps others, kill spores
by some type of inner membrane damage (Cortezzo and others 2004; Genest and
others 2002; Loshon and others 2001; Paul and others 2006, 2007; Young and Setlow
2003, 2004a,b ):
α
/
β
1. Spore populations killed 90-99% by these agents do not release their small mol-
ecule pools, in particular DPA. Thus the spore's inner membrane, the spore's main
permeability barrier to small molecules, must remain intact in the treated dormant
spore.
2. Spore populations killed 90-99% by such agents are sensitized to DPA release upon
a normally sublethal heat treatment (Cortezzo and others 2004), suggesting that the
spore's main permeability barrier to small molecules, the inner membrane, is
damaged by treatment with these agents.
3. Spore populations killed 90 - 99% by these agents exhibit much lower recovery
when plated on media with relatively high salt content in comparison to untreated
